Polyacrylamide and other acrylamide-based polymers are used as support matrices for various electrophoretic processes, including separation procedures such as isoelectric focusing, conventional zone, capillary zone, or discontinuous zonal electrophoresis. Various modifications in the pore size of the acrylamide support medium are often required for separations based on either the charge, size, or both, of macromolecules of biological origin. For example, low resolution simple charge separations of the plasma proteins may be made on media such as agarose, cellulose acetate and paper that contain pore sizes too large to sieve the molecules.
High resolution separations of macromolecules based on size require a support medium where the pore size can be varied to exert a sieving or retardation effect, on the macromolecules as they migrate in the electrical field. Such separations require a system wherein the pore size may be varied or produced with a graduated pore size. This is commonly done using polyacrylamide gels or acrylamide derivatives, where the pore size may be varied by varying the monomer concentration. An electrophoretic gel having a single pore size will not resolve many of the macromolecules present in a complex mixture.
The article Allen et al, "Rehydratable Gels: A Potential Reference Standard Support for Electrophoresing PCR-Amplified DNA" published in Biotechnology, Vol. 8, Dec. 1990 by the present Applicants has indicated that rehydratable gels can be modified after their formation by adding specific amounts of one or more alcohols, polyols or monosaccharides. The matrix modifiers were added to preformed polyacrylamide gels and were used to separate various macromolecules.
Accordingly, there is a continued need in this art for an improved method for resolving mixtures of biological macromolecules using a modified electrophoretic support to increase the resolution of macromolecules such as polynucleotides and polypeptides. Further, there is a need in this art to further improve the resolution of electrophoretic support media whose matrix has been modified to enhance the separations of macromolecules and individualization process of various biological materials.